Indirect inhibition of 26S proteasome activity in a cellular model of Huntington's disease

• Published in: The Journal of cell biology Vol. 196; no. 5; pp. 573 - 587
• Main Authors: Hipp, Mark S, Patel, Chetan N, Bersuker, Kirill, Riley, Brigit E, Kaiser, Stephen E, Shaler, Thomas A, Brandeis, Michael, Kopito, Ron R
• Format: Journal Article
• Language: English
• Published: United States Rockefeller University Press 05-03-2012; The Rockefeller University Press
• Subjects: Animals; Cell Line; Cells; Cytoplasm; Enzyme Stability; Genes, Reporter; HEK293 Cells; Humans; Huntingtin Protein; Huntington Disease - physiopathology; Huntingtons disease; Mutation; Nerve Tissue Proteins - genetics; Nerve Tissue Proteins - metabolism; Neurodegeneration; Nuclear Proteins - genetics; Nuclear Proteins - metabolism; Peptides - metabolism; Proteasome Endopeptidase Complex - metabolism; Proteasome Inhibitors; Proteins; Recombinant Fusion Proteins - genetics; Recombinant Fusion Proteins - metabolism; Ubiquitin - genetics; Ubiquitin - metabolism; Ubiquitination
• ISSN: 0021-9525; 1540-8140
• DOI: 10.1083/jcb.201110093

Pathognomonic accumulation of ubiquitin (Ub) conjugates in human neurodegenerative diseases, such as Huntington's disease, suggests that highly aggregated proteins interfere with 26S proteasome activity. In this paper, we examine possible mechanisms by which an N-terminal fragment of mutant huntingtin (htt; N-htt) inhibits 26S function. We show that ubiquitinated N-htt-whether aggregated or not-did not choke or clog the proteasome. Both Ub-dependent and Ub-independent proteasome reporters accumulated when the concentration of mutant N-htt exceeded a solubility threshold, indicating that stabilization of 26S substrates is not linked to impaired Ub conjugation. Above this solubility threshold, mutant N-htt was rapidly recruited to cytoplasmic inclusions that were initially devoid of Ub. Although synthetically polyubiquitinated N-htt competed with other Ub conjugates for access to the proteasome, the vast majority of mutant N-htt in cells was not Ub conjugated. Our data confirm that proteasomes are not directly impaired by aggregated N-terminal fragments of htt; instead, our data suggest that Ub accumulation is linked to impaired function of the cellular proteostasis network.